BacMam

The BacMam gene delivery technology is a transient expression system, which facilitates expression of toxic gene products. It has a broad range of transduction including many primary cell types and stem cells.[8] The baculoviral genome has a large capacity for foreign gene insert with up to 38 kb have been tried successfully.[9] Simultaneous delivery of multiple genes to the same cell is feasible.[10] There is little to no microscopically observable cytopathic effects of BacMam particles on mammalian cells.[11] The level of gene expression can be adjusted by viral dose or chemical additions using histone deacetylase inhibitors.[12] Transduction of cells is performed by liquid only addition and therefore BacMam is amenable to automated methods. Finally viruses are stable when stored at 4 °C in the dark for long periods of time.[13]

Baculoviruses are Risk Group 1 agents that have been widely used for over 25 years for insect cell protein production applications.[14] Baculoviruses are produced in insect cells and incapable of replicating in mammalian cells and are not known to cause disease in healthy human adults. Furthermore, BacMam viruses are inactivated by human complement, which reduces risk to researchers. Lastly, viruses used in the laboratory cannot replicate in insects so there is no environmental threat from these particles accidentally being released into the environment.[15][16]

Studies on baculovirus entry into human hepatocellular carcinoma cells suggest that BacMam enters mammalian cells via clathrin-mediated endocytosis and possibly via macropinocytosis.[17] Further studies have suggested that caveolae are somehow involved in baculovirus entry in mammalian cells.[18]

To be effective, a gene delivery technology must not interfere with normal cellular function. Cytotoxicity assays and transcriptome analyses on a human HEK cell line (HEK293) have revealed that baculovirus transduction is not cytotoxic and does not induce differential transcriptional responses.[19] Similarly, infected Schwann cells retain their characteristic morphological and molecular phenotype and are capable of differentiating in vitro and express the P0 myelination marker. Using complementary DNA (cDNA) microarray technology to examine in vitro and in vivo global cellular gene expression profiles in the rat brain, cultured human astrocytes and human neuronal cells after viral transduction, host antiviral responses were observed.[20] The related genes were mainly those associated with innate immunity, including several of the genes involved in Toll-like receptor signaling pathway and cytokine-cytokine receptor interaction.

• Bioproduction

BacMam has been used to produce proteins in large quantities using HEK293 cells in a hollow fiber bioreactor system[21]

• High Throughput Screening

Pharmacology of G protein-coupled receptor is enabled with the use of BacMam technology in drug discovery applications[22]

• Fluorescence Microscopy

Organelle labeling reagents are commercially available BacMam particles for labeling organelles and other subcellular structures[23]

Single mitochondrion labeled with a mitochondrial targeted green fluorescent protein[24]

• Receptor Activation/Pathway Analysis

Characterization of serotonin receptor activation via a BacMam delivered GFP fusion to a kinase substrate[25]

• Structural Biology

BacMam system has been used to produce soluble and membrane glycoproteins for structural studies[26]

• Baculovirus

• The BacMam System